My long-term goal is to understand how cells maintain genomic stability and how failure to do so results in the genetic instabilities associated with cancer and human disease. To achieve these goals, I will establish myself as independent investigator in my own lab to continue my studies of Pif1p and telomerase in the yeast Saccharomyces cerevisiae. My success will positively impact minority representation in the biomedical sciences by establishing a research program to train talented students. Telomeres are the specialized functional structures found at the ends of linear eukaryotic chromosomes. Telomeres are essential for the stable maintenance of linear chromosomes because they protect the chromosome ends from DNA degradation. This proposal focuses on Pif1p, a conserved 5'to 3'helicase that inhibits telomerase in vitro and in the cell. The specific aims of this proposal are designed to further probe the role of Pif1p in protecting the genome from DNA damage. Pif1p helicase is a cell cycle regulated protein. However, the role of Pif1p modifications, regulation and protein stability on Pif1p function and/or activity is currently unknown. The central hypothesis that underlies the aims of this proposal is that Pif1p levels are tightly regulated and that inappropriate expression/ activity of Pif1p results in DNA damage and genomic instability. Consistent with the hypothesis, I found that over-expression of Pif1p throughout the cell cycle results in loss of viability in strains with defective telomere end protection. In the first aim I will conduct experiments to examine the cell cycle regulation and protein modifications of Pif1p. As part of this aim, I will determine the effects of DNA damage on Pif1p and I will test the role Piflp in the meiotic cell cycle. My hypothesis suggests that Pif1p levels may be critical in certain circumstances -- for example, after DNA damage or in combination with other mutations. By direct test, I identified such a condition, Pif1p over-expression is lethal in strains compromised for telomere end protection including cdc13-1 and yKU mutants. I will use Pif1p's over-expression lethality as the basis for genetic screens to identify genes that affect telomerase and telomere end protection. Since many of the components involved in telomere maintenance and DNA metabolism are highly conserved between yeast and higher eukaryotes, the successful completion of these studies may give insight into of the mechanisms that lead to the genetic instabilities associated with cancer.